The effects of parathyroid hormone (PTH) and prostaglandin E2 (PGE2) on glycosaminoglycan (GAG) synthesis in bovine dental pulp cells were studied. Dibutyryl cyclic adenosine 3',5'-monophosphate and isobutyl methylxanthine were used to assess whether their effects were mediated by intracellular cAMP. Glycosaminoglycan synthesis was assayed by measuring [35S]sulfate incorporation into the GAG fraction of dental pulp cells. Glycosaminoglycan synthesis was increased 1.3-fold by PTH (4 units per ml) alone, 1.6-fold by PTH in the presence of isobutyl methylxanthine, 1.2-fold by PGE2 (100 ng per ml) alone, and 1.5-fold by PGE2 in the presence of isobutyl methylxanthine. Dibutyryl cyclic adenosine 3',5'-monophosphate enhanced GAG synthesis in a concentration-dependent manner and mimicked the effects of PTH and PGE2. The effects of these hormones on pulp and gingival cells were compared; addition of PTH, PGE2, and dibutyryl cAMP had no effect on gingival cell GAG synthesis, whereas their addition induced significant increases of GAG in pulp cells. These results indicate that PTH and PGE2 are involved in the differentiation of dental pulp cells and that these effects are mediated by cAMP.